Q: Can you provide some historical perspective on the application and development of mass spectrometry (MS) techniques and instruments for clinical research?
A: In the clinical world, gas chromatography-MS has been in use for a while, but it’s the advent of the triple quadrupole tandem MS that provided improved sensitivity and selectivity for analysis. The improvements had a big impact on clinical applications, such as for diagnosing a disease or finding biomarkers for research. It also enabled multiplexing, which helped researchers in this field monitor several analytes at the same time. For instance, more than 30 biomolecules or metabolites could be measured in a rapid screening, which becomes important when diagnosing many diseases in one sample for applications like newborn screening. The coupling of MS and chromatography with electrospray ionization made it easier to analyze almost any type of molecule, including nonvolatile samples, and this also helped us diagnose many rare diseases during newborn screening. There was really no limit in terms of the types of biomolecules that could be measured using this technique. The only limitation was to get the right sample to find out what that signal meant in a biological context.
Q: Can you provide details about your work in developing instrumentation and applications for imaging MS studies?
A: As an undergraduate in analytical chemistry, I started using matrix-assisted laser desorption/ionization (MALDI)-MS to look at protein signatures in bacteria and then, during my graduate work, developed MS-based imaging primarily for studying small molecules in tissue. Using the capabilities of tandem MS with an ion trap, we could identify and characterize the molecules, and imaging was used to look at their tissue distributions. Now as a faculty member in the department of pathology, I use a lot of those same approaches for clinical research. One of the advantages of imaging MS is that you can directly take a tissue sample and analyze what is present, which helps you avoid sample loss or changes in the analytes during an extraction. We are developing quantitative MS techniques that will enable us to look at target distribution in tissue that may be missed using extractive approaches.
MALDI enables you to identify a wide range of analytes, from small molecules and peptides to large proteins and biomolecules. As long as you can ionize it, you can analyze it. This has definitely helped quantitative analysis. However, one of the limitations of MALDI is that you have to coat the tissue with the matrix to get it ready for analysis. Hence, we have developed a spray chamber to coat multiple tissues with the matrix in a very rapid way to enable high-throughput studies. We are also working on developing techniques to avoid using MALDI and not have to coat the tissues. One of these techniques is the liquid microjunction surface analysis that we use to understand rapid changes in metabolites associated with Parkinson’s disease (PD). Here, we keep the tissue in a medium so that it remains in an active state and then rapidly analyze the changes that are taking place after electrical stimulation, a current treatment for PD.
We are also doing a lot of work related to understanding the microbiome. In one of the studies, we are using extractive liquid chromatography-MS to study the microbes that either inhibit or enhance tryptophan-mediated metabolism in a rodent model. The goal is to understand how the microbiome impacts tryptophan metabolism, which in turn affects various neurotransmitters and changes brain chemistry. Eventually we would like to use imaging MS to look at the distribution of the microbiome and study how the changes in distribution affect brain development.
Q: What are the existing challenges and gaps in the use of MS in the clinic?
A: One of the biggest challenges in using imaging MS is still its throughput. An MS scan takes anywhere between 30 minutes to two hours and can take even longer depending on the size of the sample. There are new instruments under development that we’re using to try to speed up this process, but it’s still not as fast as looking under a microscope at a histology stain to identify what is present. Hence, developing faster imaging-based approaches for analysis will certainly help, but more important is finding ways to figure out whether imaging is really needed or not. The value of imaging, however, lies in the amount of information you can gather from each scan to help better understand the biological mechanisms. Imaging MS is information rich, containing several hundred molecules in a single mass spectrum.
It will take some time to translate imaging MS to routine diagnostic applications, and we may have to start with using it for cases where we have a problem using histology to make a diagnosis. Histology has been used for many years and has a large repository and knowledge base. Hence, one of the biggest advantages of histology is being able to go back and pull a tissue sample from years ago to compare it to the one you have today. Imaging approaches can also benefit from having such databases where you can compare tissue samples from the past to a scan that was recently generated. Creating data repositories for labs that do imaging MS will certainly help. Imaging MS is used in tandem with histology on tissue biopsies. With histology, you target a single analyte or a certain class of molecules, whereas with MS you can look at individual chemical signatures for many different analytes in the tissue. This gives a better understanding of the biology, which can help you understand different manifestations of the same disease and can lead to better treatment options. The other big trends in the field today are coupling imaging MS with NMR or MRI and developing stains or agents that will target new biomarkers.
Q: Any advice to our readers who are looking to explore the use of imaging MS?
A: One of the key things is finding opportunities to get adequately trained in using imaging MS through workshops at conferences like American Society for Mass Spectrometry (ASMS) or the Imaging Mass Spectrometry meeting. At these meetings, you can also learn new approaches that key leaders in the field are applying and make connections with people whom you can reach out to for assistance. Hiring people who already have the necessary experience will certainly help. Vendors have done an excellent job of developing the instrumentation, as well as the tools to analyze the data. Hence, reaching out to them to understand what the instruments are capable of and providing them with samples to analyze can help you see the possibilities that exist. With imaging MS, you get so much data on the chemical signatures that trying to figure out which chemical species is the most relevant to the question you are asking is very important. Finding the right software tools to help with data analysis and developing quality control protocols to make sure that a sample is handled in the same way, every time, ensures that the data is reliable.
As an MS consultant, I find that some of the common problems are associated with finding out when the instrument is not working properly. Helping people find ways to quickly figure out whether the problem is with the instrumentation, with sample preparation, or with different components along the way, becomes important. How to maintain the quality data or the analysis is also of concern to most people. When starting a new lab, there are always questions around how to routinely maintain the MS or how to build a lab that enables you to effectively use MS. It’s often the little details that make a big difference.
Timothy Garrett received his undergraduate degree in chemistry from the University of Georgia, graduating summa cum laude. As an undergraduate, he worked in the lab of Dr. I. Jonathan Amster on the characterization of bacterial proteins using MALDI-TOF. After two years in industry, he enrolled in the PhD program at the University of Florida, working under the direction of Dr. Richard A. Yost. As a graduate student, he worked on the first imaging mass spectrometry-based ion trap instrument through a partnership with Thermo and studied the disposition of phospholipids in brain tissue. He is currently an associate professor in the Department of Pathology at the University of Florida, where he is associate director of the Southeast Center for Integrated Metabolomics (SECIM). His current interests are the application of direct tissue analysis approaches such as MALDI, DESI, and LMJ-SSP, as well as the use of high-resolution mass spectrometry in metabolomics and routine diagnostics. He enjoys the interplay between technological advancement and clinical analysis, believing they provide unique opportunities to develop future diagnostic tools.